The broad goal of this research program continues to be the definition of the mechanisms by which acid hydrolases are transported to lysosomes. This involves delineation of the receptors which mediate intracellular sorting of lysosomal enzymes and uptake of lysosomal enzymes by adsorptive pinocytosis, and delineation of the recognition mark on the enzymes through which they act as ligands for these receptors. It also involves studies of the processes of vesicular transport that deliver newly synthesized enzyme from the endoplasmic reticulum to lysosomes, and endocytosed enzyme from the plasma membrane to lysosomes. Specific aims include: 1) Characterize the oligosaccharides on human spleen Beta-glucuronidase that bind to the phosphomannosyl receptor (PMR) and mediate enzyme transport. 2) Determine the basis by which Beta-glucuronidase and Beta-hexosaminidase B are recognized by the targeting enzyme, N-acetylglucosaminyl phosphotransferase. 3) Determine the kinetics of inactivation of the recognition marker following transfer of enzyme to lysosomes. 4) Determine the reasons for lysosomal enzyme secretion by normal fibroblasts. 5) Compare the turnover of metabolically labeled, newly synthesized PMR with that of cell surface PMR labeled with lactoperoxidase. 6) Determine whether endocytosed acid hydrolases enter the Golgi apparatus for sorting enroute to lysosomes. 7) Study the transport of acid hydrolases and of myeloperoxidase in leukocytes, and in the human leukemic cell line HL60. 8) Define the role of the "proton pump" in transport of lysosomal enzymes, by analysis of mutants defective in acidification of lysosomes. The answers sought have fundamental significance and will help define the requirements for enzyme replacement for inherited lysosomal storage diseases.